Orientation-Controllable Enzyme Cascade on Electrode for Bioelectrocatalytic Chain Reaction

Area of Science:

• Bioelectrochemistry
• Enzyme Engineering
• Nanobiotechnology

Background:

• Achieving efficient direct electric communication in multienzyme electrodes is complex due to intricate binding requirements.
• Coimmobilizing enzymes for cascade reactions on electrode surfaces presents challenges in maintaining optimal conformation and activity.

Purpose of the Study:

• To construct an enzyme cascade-induced bioelectrocatalytic system using solid binding peptide (SBP) for coimmobilizing invertase (INV) and flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase gamma-alpha complex (GDHγα).
• To investigate the impact of interenzyme orientation and binding conformation on direct electron transfer (DET) and overall cascade efficiency.
• To demonstrate the utility of SBP fusion technology for developing generic cascade-induced direct bioelectrocatalytic systems.

Main Methods:

• Engineered a fusion enzyme cascade system by coimmobilizing INV and GDHγα on an electrode surface using SBP as a molecular binder.
• Strategically designed the SBP-fused enzyme cascade to control relative enzyme orientations and facilitate direct electron transfer (DET) at the FAD cofactor of GDHγα.
• Analyzed how interenzyme orientation influences intermediate delivery and overall chain reaction efficiency.

Main Results:

• Demonstrated that the interenzyme relative orientation significantly impacts the intermediate delivery route and cascade efficiency.
• Showed that interfacial DET between the fusion GDHγα and the electrode is modulated by the binding conformation of coimmobilized enzymes.
• Validated the efficacy of SBP fusion technology in creating well-defined enzyme orientations for enhanced bioelectrocatalysis.

Conclusions:

• Emphasized the critical role of interenzyme orientation in enzymatic cascade systems for electrocatalysis.
• Highlighted SBP fusion technology as a versatile tool for constructing cascade-induced direct bioelectrocatalytic systems.
• Indicated the broad applicability of this approach in enzyme cascade-based bioelectronics, including biofuel cells, biosensors, and bioelectrosynthetic systems.